Mitochondria are the primary energy source of most eukaryotic cells. Each mitochondrion possesses multiple copies of mitochondrial DNA (mtDNA). The human mitochondrial genome is a closed circular molecule of DNA 16 kb long. It encodes genes for 13 electron transport chain proteins, 22 tRNAs, and two rRNAs. The mitochondrial genome also includes a control region that contains the displacement loop (D-Loop), within which DNA replication is initiated and gene transcription is regulated. By convention, one particular sequence, known as the revised Cambridge Reference Sequence (rCRS), or the Anderson sequence, serves as a reference sequence to which the other sequences are compared (Anderson et al. (1981) Nature 290, 457-465.; Andrews et al. (1999) Nature Genetics 23, 147.; herein incorporated by reference in their entireties). Recently, increased levels of mutations within the mitochondrial genome have been linked to several diseases, including diabetes, Alzheimer's, and cancer (Wallace (1994) Proceedings of the National Academy of Science, 91, 8739-8746.; Pravenec et al. (2007) Genome Research, 17, 1319-1326.; Swerdlow (2004) Medical Hypotheses, 63, 8-20.; Chen et al. (2004) Journal of Environmental Science and Health, C22. pp. 1-12.; herein incorporated by reference in their entireties). Despite these insights a substantial technical challenge persists in the areas of detecting, characterizing, and diagnosing changes in mtDNA sequence largely because there are very large numbers, tens to many thousands, of mtDNA molecules in each eukaryotic cell. This means that changes in mtDNA molecules are often “averaged out” in populations of mtDNA molecules, even in single cells. What is needed are better systems and methods for characterizing mtDNA to assist in biological research, drug development, assessment and monitoring of drug or therapeutic impact, and disease screening, diagnosis, and monitoring.